Determining the mechanisms that allow neoplastic cells to undergo uncontrolled proliferation is crucial to understanding carcinogenesis. Recent studies have made it very clear that protein phosphatases, like protein kinases, play important and specific roles in cell-cycle regulation; thus alterations in the expression or regulation of these proteins may affect cell-cycle progression and could even allow cells to undergo unregulated proliferation. We have cloned and characterized four human PPases, and one, designated PP5, is particularly attractive for further studies. PP5 acts to suppress a glucocorticoid-induced, p53- mediated signaling cascade leading to the induction of G1/S-phase growth arrest. Studies into the mechanisms regulating PP5 activity have revealed that the expression of PP5 is responsive to 17-13 estradiol and hypoxia inducible factor-1 (HIF-1), which are both positive factors in the development of human breast cancer. Furthermore, the constitutive over expression of PP5 aids cell survival during oxidative stress and converts MCF-7 cells from an estrogen-dependent into an estrogen-independent phenotype. Thus, aberrant PP5 activity may also contribute to tumor development. This proposal is designed to test the hypothesis that protein phosphatases play an important role in cell cycle progression; thus, abnormal expression or the interference of their normal activity may contribute to the aberrant proliferative behavior of neoplastic cells. These studies will focus on the following specific aims: Aim 1. Continue to characterize the roles played by PP5 in glucocorticoid receptor-mediated signaling networks. Aim 2. Determine the roles of PP5 in HIF-1- and estrogen-mediated signaling cascades that aid cell survival and in cross-talk between p53-, GR- and HIF-1-mediated signaling networks. Aim 3. Determine if PP5 expression is a positive factor in the development of human breast cancer, and characterize the physiological role for PP5 through the development of PP5-knockout and PP5-loxP transgenic mice. Through these studies we hope to further characterize the role of protein phosphatases in the regulation of cell cycle control and gain insight into the how protein phosphatases may be involved in the aberrant proliferation of neoplastic cells.